Gene synthesis is integral to molecular biology, generating probes for life science industries. Such industries include drug discovery, vaccine development, gene therapy, tissue replacement, human genomics, disease detection research, and improving the nutritional quality of food.
Classic genetic methods used a few decades ago only facilitated the modification of one molecule at a time. However, the next-generation sequencing services provided by gene synthesis companies now facilitate the production of recombinant genes, mutated genes, and novel DNA via custom gene synthesis.
Gene synthesis is the foundation for related services like peptide synthesis, protein production, antibody production and sequencing, and ELISA kit production. Learn more below.
What Is Gene Synthesis?
A gene is the primary genetic unit carrying genetic information or hereditary traits transferable from parent to offspring. It features nucleotides that bind together to form a DNA molecule. Genes occur naturally as nucleic acid sequences in the chromosomes present in the human cell. However, gene synthesis creates synthetic gene material nucleic acid sequences as base pairs.
Gene synthesis services allow researchers to optimize or modify genetic components to boost the target gene’s traits or characteristics for use as probes in various life sciences research projects.
Traditional gene synthesis techniques included mutagenesis, or introducing a mutagen (mutation-causing agent) in a genetic code. However, mutagenesis only altered single base pairs in an entire gene sequence and, as such, was not viable for projects like custom DNA synthesis or custom gene synthesis.
The second classic gene synthesis option was traditional cloning with restriction enzymes. A restriction enzyme is a bacteria-generated protein that cleaves or cuts DNA sequences along target sections to create DNA fragments like genes.
While traditional cloning has multiple advantages, including low labor intensity and diverse expression systems, such as the E coli protein expression system that DNA synthesis companies use in providing protein expression and purification services, the technique also has limitations.
Classical custom gene synthesis techniques had disadvantages, including possible sequence constraints resulting from restriction sites. Other disadvantages included undesirable recombination sequences in protein production services and a vector structure that could affect downstream services like recombinant protein production and antibody sequencing services.
The first historical mention of gene synthesis was in 1972, when H. Gobind Khorana, an Indian biochemist, developed the first in vitro functional synthetic gene. He used multiple enzymes to link DNA fragments into a gene.
Fast-forward to the present day, and advancements in biotechnology make it possible to generate an entire synthetic genome from gene synthesis, create difficult-to-extract natural gene sequences, and develop novel sequences that do not occur naturally.
Most gene synthesis companies offer custom DNA synthesis services and can generate 150–200 kilobase pair (kb) synthetic gene lengths using techniques like the solid-phase peptide synthesis protocol. Moreover, some gene synthesis companies use ligation techniques to generate longer DNA strands with thousands of base pairs.
Therefore, besides transferring genetic traits to offspring, the genes in DNA molecules are vital components in peptide synthesis and protein production services like antibody production. Consequently, gene synthesis is a foundational product for gene synthesis companies.
However, such companies build on gene synthesis services by offering related services. Such services include peptide synthesis, custom protein synthesis, custom antibody services, custom antibody expression, protein expression services, antibody sequencing services, and ELISA kit production.
Custom gene synthesis companies have varying costs depending on the length and complexity of the sequence, expression systems, and the level of customization. Other DNA synthesis companies charge clients per base pair.
Nonetheless, gene synthesis has revolutionized biotech processes, including protein production services, protein expression services, solid-phase peptide synthesis protocols, antibody production, and peptide synthesis services. Below is an overview of the benefits of gene synthesis to DNA synthesis companies and recombinant protein production services in the life sciences.
Gene Synthesis Advantages
In vitro gene synthesis services have multiple advantages over traditional cloning techniques. Below are the top reasons why DNA synthesis companies offer gene synthesis services as a primary product.
1. Quick Turnaround Time
Classic gene synthesis methods had a slow turnaround time, taking service providers months to deliver the target gene required for research and industrial purposes. As a result, research projects would stall, and others would shut down altogether due to the lengthy timelines it took DNA synthesis companies to generate genes and DNA products.
In contrast, contemporary gene synthesis methods, like modern solid-phase synthesis protocols, have a significantly shorter turnaround time. Although the turnaround time varies depending on the gene length and subcloning difficulty, the industry average turnaround time for gene fragments is 6–9 days, while the turnaround time for clonal genes is 14–21 days.
Moreover, some gene synthesis providers offer complimentary services, including free standard vectors for subcloning in protein expression systems. Additionally, the final gene product delivery as a lyophilized plasmid takes less than 24 hours within US borders. Therefore, researchers can order gene products online easily from DNA synthesis companies.
2. High Accuracy Levels
Most gene and custom DNA synthesis service providers guarantee 100% gene sequence accuracy. Custom DNA synthesis companies use PCR (polymerase chain reaction) methods to ensure gene product accuracy.
PCR is the standard method for amplifying exact copies of a target DNA or gene molecule into millions of copies. High accuracy in gene replication is crucial to ensure consistent research results or product delivery in applications like antibody production, protein expression, recombinant protein production, ELISA kit production, and custom protein production.
Although multiple PCR methods are available, dial-out PCR is the standard industry technique for ensuring gene product accuracy. Moreover, some gene delivery services provide complete gene sequencing quality control data. Such data includes a quality analysis report, a construct map, the entire sequence, chromatograms, and an alignment file.
3. Affordability
The increasing availability of gene synthesis services makes gene product services like peptide synthesis, custom protein synthesis, custom antibody production, and protein expression and purification services more affordable. Moreover, most service providers have a "no gene, no charge" policy, protecting researchers and commercial biotech stakeholders from potential losses arising from low-quality genes or other DNA products.
Gene Synthesis Protocol / Standard Workflow
Gene synthesis is a step-by-step process that begins with an oligonucleotide (a short DNA or RNA fragment) and ends with a gene plasmid ready for delivery and use in an industrial or lab setting.
The two primary methods used in gene synthesis are polymerase cycling assembly (PCA) and gene synthesis by ligation. Although each method has advantages and disadvantages, both techniques follow a similar protocol to maximize the target gene yield. Below is an overview of the gene synthesis protocol leading to a high-quality, active gene molecule.
1. Chemical Synthesis of Oligonucleotides
Gene synthesis begins with the chemical synthesis of oligonucleotides. This step is crucial for recombinant protein production processes like custom antibody services, custom protein synthesis, custom protein expression, peptide synthesis services, and custom protein production.
Chemical oligonucleotide synthesis occurs in the 3’ to 5’ direction and entails using a solid-phase synthesis protocol to attach a chemically-modified nucleoside to a polymeric support. The polymeric support holds a growing oligonucleotide chain, which undergoes deprotection, similar to the processes used in recombinant protein production and peptide synthesis services.
2. Oligonucleotide Annealing By Ligation or Polymerase Reactions
The newly-formed oligonucleotides undergo annealing as the second step in the gene synthesis protocol. Annealing entails heating the molecules and then cooling them slowly to facilitate hybridization. In this context, hybridization refers to the binding of complementary DNA strands.
As earlier stated, an oligonucleotide is a DNA fragment and features the double-strand structure typical of DNA molecules. Therefore, the heating process in annealing splits the double-stranded oligonucleotide into single strands.
On the other hand, the cooling process merges multiple single-strand oligonucleotides into a single long chain. However, care is necessary to prevent E. coli protein aggregation or property alteration during annealing.
PCA and synthesis by ligation are the primary processes used in oligonucleotide annealing. Custom protein production processes like custom antibody production and custom protein expression dictate the ideal annealing method.
3. Gene Sequence Cloning Into Vectors
Cloning is a process entailing the replication of pieces of DNA into multiple copies. Oligonucleotide annealing facilitates the formation of a complete gene sequence. Therefore, the next step is cloning that gene for research and industrial purposes like peptide synthesis services, custom antibody services, protein expression services, and recombinant protein production.
A cloning or plasmid vector is a piece of DNA that replicates independently from the host cell and is the primary replication machinery in gene synthesis. Although natural cloning vectors like E. coli protein expression systems are available, synthetic plasmids often require fewer steps for protein purification. Therefore, such plasmids are often better options for protein expression and purification services.
4. Clone Screening
Clone screening is necessary to identify and isolate acceptable target genes from the E. coli protein expression system. While ELISA (Enzyme-Linked Immunosorbent Assay) kits are ideal clone screening tools for detecting the target protein in a biological sample, they are frequently used alongside other methods.
Gene product service providers typically partner with ELISA kit manufacturers to acquire assays for effective clone screening. However, other screening methods, like chromatography, are available in addition to test kits from ELISA kit manufacturers.
5. Sequencing Analysis
Gene sequencing entails determining the genetic composition of a gene or DNA by examining the base pairs. The process utilizes sequencing primers, including gene-specific primers, to reveal the exact gene sequence.
Additionally, gene sequencing goes hand-in-hand with gene analysis and functional studies. Gene product research studies on antibody production services, antibody specificity, monoclonal antibody sequencing, and polyclonal antibody production can be conducted using sequenced genes retrieved from E. coli protein expression systems.
6. Error Correction
Replication errors like base substitution, deletion, and incorrect insertions are common during gene assembly. Although such errors negatively impact analysis processes like antibody specificity and monoclonal antibody sequencing, error correction is performed to ensure the accuracy of the final construct.
7. Plasmid Purification and Lyophilization
Different research projects, such as antibody production and antibody sequencing services, require different gene purity levels. While synthetic genes require efficient purification, highly automated workflows mean that standard purification is often included. Moreover, some service providers offer advanced protein purification as a value-added service.
Gene Synthesis Applications In Biotechnology
As earlier stated, gene synthesis is a foundational process for most biotech applications. Below are the top gene synthesis applications for research and industry.
1. Antibody Production
Antibody production entails the introduction of antigens into a host to generate target antibodies. Although antibody production is a natural process, gene synthesis offers multiple advantages, including preventing side reactions and toxicity to the host. Additionally, synthetic genes have enhanced properties, including higher antibody specificity and a lower likelihood of E. coli protein aggregation.
Custom antibody services, monoclonal antibody sequencing services, and the steps to produce polyclonal antibodies begin with gene synthesis. Moreover, gene sequencing lowers the cost of monoclonal antibody sequencing. Research into polyclonal antibody production has also contributed to developing potent antibodies with multiple binding sites for enhanced specificity.
2. Recombinant Protein Production
Recombinant protein production is a multifaceted process featuring elements such as protein expression services, custom protein production, antibody sequencing, and custom antibody production.
Gene synthesis is the starting point for the above processes, facilitating solid-phase techniques and the steps for protein purification. Additionally, recombinant protein production utilizes high-throughput techniques to generate high-quality proteins for therapeutic drug discovery and genetic modification for food security. According to one report, over 200 FDA-approved therapeutic proteins and peptides are currently available for clinical use in the U.S.
3. ELISA Assay Test Kit Production
An Enzyme-Linked Immunosorbent Assay (ELISA) kit is an assay tool featuring antigens or antibodies embedded on multi-well plates. So, how does ELISA work? ELISA kit manufacturers design the kits with antibodies to measure the presence and volume of targeted proteins.
The kits have multiple applications in disease detection and diagnosis. Also, ELISA kit manufacturers optimize the kits for protein specificity during the development process. Most gene product service providers incorporate the ELISA test kit cost into their research packages, lowering overall custom antibody costs.
Conclusion
Gene synthesis has revolutionized biotechnology by facilitating research to develop optimized genes that improve the quality of life within and beyond medical fields. Synthetic genes enable high-throughput gene product generation for the agricultural, pharmaceutical, beauty, waste treatment, and food processing industries. Therefore, gene synthesis is the future of biotechnology, and researchers are constantly working on new methods to make the process more cost-effective and efficient.